Project Summary/Abstract Inheritance of parental traits by offspring is the most fundamental process for the perpetuation of life on earth and is vital for the process of evolution. Although there is mounting evidence from worms to humans suggesting that parental environment can influence phenotypes in offspring, the mechanism of such transgenerational inheritance ?sometimes referred to as inheritance of acquired traits, or Lamarckian Inheritance? remains deeply mysterious. As epigenetic marks are responsive to environmental conditions, intergenerational inheritance is likely mediated via alterations to the gamete ?epigenome?. This project aims to elucidate how environmental conditions modulate specific epigenetic marks in germ cells and how those marks influence the development of offspring. The discovery that our environment can affect the phenotype of future generations has tremendous significance for basic biology and for public health and policy. Male mice fed a low protein diet sire offspring with altered lipid and cholesterol metabolism. My previous studies revealed that paternal dietary information is transmitted to offspring via sperm and that small RNAs are the environmentally- responsive epigenetic molecules in sperm. Surprisingly, I found that RNAs in mature sperm are synthesized in the somatic cells of the epididymis a long-convoluted tubule where sperm undergo post-testicular maturation and shipped to sperm via extracellular vesicles. These astonishing observations suggest that small RNA-mediated communication between the somatic cells and the maturing gametes is a potential mechanism for signaling paternal environmental information to offspring, and provide a focus for this proposal to investigate the mechanism of RNA-mediated intergenerational inheritance by elucidating four key steps of the process: 1) how parental somatic cells generate informational signals for gametes, 2) how those signals are influenced by environment, 3) how those signals are transmitted to gametes, and 4) how those signals influence offspring gene expression and development. I will elucidate these steps using a unique and powerful combination of molecular, genetic, reproductive, and genomic approaches in mouse as a mammalian model. The proposed studies are high-impact with a potential to shift paradigms, as they aim to investigate the role of a somatic tissue (epididymis) in shaping gamete epigenome and influencing inheritance. Moreover, these studies will have a tremendous impact on the fields of RNA biology, reproduction, vesicle trafficking, development, evolution, and epidemiology.